Multiomics Analyses Reveal Sex Differences in Mouse Renal Proximal Tubules

Sex differences have been increasingly recognized to play an important role in kidney physiology and pathophysiology, but limited resources are available for comprehensive interrogation of sex differences. Here, we coupled renal tubule microdissection with RNA-seq and ATAC-seq with a focus on proximal tubules that exhibited the greatest sex differences and carried out whole-kidney proteomics to build a comprehensive sex difference landscape between male and female mouse kidneys. The transcriptomic data indicate that the major sex differences are in S2 and S3 of proximal tubule segments, where signaling pathways including monocarboxylic acid metabolic process, organic anion transport, and organic acid transport display the greatest differences. We develop an ATAC-seq method on microdissected tubules to capture chromatin accessibility differences between sexes. The analysis reveals that major sex differences are in autosomes instead of sex chromosomes, and many of them are in S2/S3 segments related to the signaling pathways identified by transcriptomic analysis. Motif analysis identifies several transcription factors (Tead1, Nfia/b, and Pou3f3) whose interplay with other transcription factors (e.g. Hnf1b, Hnf4a) may contribute to sex differences. Finally, whole-kidney quantitative proteomics analysis correlates with the transcriptome analysis and identifies a large number of proteins (Cyp2e1, Acsm2/3) that are sex-dependent. We develop a knowledge portal to promote our understanding of sex differences in kidneys in kidneys at https://esbl.nhlbi.nih.gov/MRECA/PT/. Here, we combined renal tubule microdissection with RNA-seq and ATAC-seq to characterize gene expression and chromatin accessibly in proximal tubule S1, S2, and S3. We provide insights into specific genes and pathways that differ between male and female mouse kidneys. We identify sex-dependent cis-regulatory elements and reveal the interplay between transcription factors, which may result in sex-biased gene expression. We report sex differences in protein expression in kidneys to complement our gene expression data. Finally, we develop a web-based portal that allows users to download and promote understanding of the sex differences in kidneys.性别差异在肾脏生理学与病理生理学中的重要作用已日益得到学界认可，但目前用于全面探究性别差异的研究资源仍较为匮乏。本研究将肾小管显微切割技术与RNA测序（RNA-seq）及转座酶可及性测序（ATAC-seq）相结合，重点聚焦于性别差异最为显著的近端肾小管，并通过全肾蛋白质组学分析，构建了雌雄小鼠肾脏之间的全面性别差异图谱。转录组数据显示，主要的性别差异集中于近端肾小管节段的S2和S3亚段，其中单羧酸代谢过程、有机阴离子转运以及有机酸转运等信号通路呈现最为显著的差异。我们针对显微切割的肾小管建立了ATAC-seq分析方法，以捕捉不同性别间的染色质可及性差异。分析结果表明，主要的性别差异存在于常染色体而非性染色体，且其中许多差异位于与转录组分析所鉴定信号通路相关的S2/S3亚段。基序分析鉴定出若干转录因子（Tead1、Nfia/b及Pou3f3），它们与其他转录因子（如Hnf1b、Hnf4a）的相互作用可能是性别差异产生的潜在机制。最后，全肾定量蛋白质组学分析与转录组分析结果相互印证，鉴定出大量性别依赖性蛋白（如Cyp2e1、Acsm2/3）。我们搭建了一个在线知识门户（https://esbl.nhlbi.nih.gov/MRECA/PT/），以增进学界对肾脏性别差异的理解。本研究结合肾小管显微切割技术与RNA-seq及ATAC-seq，对近端肾小管S1、S2及S3亚段的基因表达与染色质可及性进行了表征。我们揭示了雌雄小鼠肾脏中存在差异的特定基因与通路，鉴定出性别依赖性顺式调控元件，并阐明了转录因子之间的相互作用，这可能是导致性别偏向性基因表达的原因。我们还报道了肾脏中蛋白质表达的性别差异，以补充基因表达数据。最后，我们开发了一个基于网页的门户，允许用户下载相关数据并加深对肾脏性别差异的理解。